[Detection of breast cancer stem cells and the expression of key molecules in Hedgehog signaling pathway].
OBJECTIVE
To isolate and identify breast cancer initiating cells (CD44(+)/CD24(-/low) cells) in breast cancer cell lines MCF-7 and MDA-MB-231, and to evaluate the activity of the Hedgehog signaling pathway in different subpopulations.


METHODS
CD44+/CD24(-/low) subpopulation and non-CD44+/CD24(-/low) subpopulation from breast cancer cell lines of MCF-7 and MDA-MB-231 were separated by fluorescence-activated cell sorting (FACS). Laser Scanning Confocal Microscope was used to identify CD44+/CD24(-/low) cells. Reverse transcription-polymerase chain reaction (RT-PCR) was employed to analyze the expression of Human Patched gene (PTCH), Sonic Hedgehog (SHH), glioma-associated oncogene homoglog-1(GIi-1), smoothened homolog (SMOH) and GAPDH of CD44+/CD24(-/low) subpopulation and non-CD44+/CD24(-/low) subpopulation from MCF-7 and MDA-MB-231.


RESULTS
The CD44+/CD24(-/low) subpopulation in MCF-7 and MDA-MB-231 cell lines were (1.70+/-1.43)% and (94.2+/-1.2)% respectively. The Hedgehog signaling pathways were active in CD44+/CD24(-/low) subpopulation of MCF-7 and MDA-MB-231 cell lines. The expression of transcription factor GIi-1, which was downstream components of the hedgehog pathway, was assayed and the results showed that the level of GIi-1 mRNA of CD44+/CD24(-/low) cells was much higher than that of non-CD44+/CD24(-/low) cells (P = 0.007 in MCF-7, and 0.005 in MDA-MB-231).


CONCLUSIONS
Breast cancer initiating cells (CD44+/CD24(-/low) subpopulation) exist in MCF-7 and MDA-MB-231 cell lines. The Hedgehog signaling pathway is active in the subpopulation of MCF-7 and MDA-MB-231 cell lines.
Lu
Zhi-hao
ZH
Department of Medical Oncology, Beijing Cancer Hospital & Institute, Peking University School of Oncology, Beijing 100142, China.
Jia
Jun
J
Ren
Jun
J
Ma
Bo
B
DI
Li-jun
LJ
Song
Guo-hong
GH
D001943:Breast Neoplasms / Q000378:metabolism* / Q000473:pathology; D051927:CD24 Antigen / Q000378:metabolism; D045744:Cell Line, Tumor; D005260:Female; D053823:Hedgehog Proteins / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D018960:Hyaluronan Receptors / Q000378:metabolism; D014411:Neoplastic Stem Cells / Q000378:metabolism*; D015398:Signal Transduction
D051927:CD24 Antigen; D053823:Hedgehog Proteins; D018960:Hyaluronan Receptors